We will evaluatethe potential utility of polyamine depletion as a means of chemosensitizing tumor cells to the cytocidal effects of anti-neoplastic drugs. Partial polyamine depletion will be achieved by incubating cultured human tumor cells with difluoromethylornithine (DFMO). Using colony forming efficiency assays to measure cell survival, we will compare dose-response curves to three alkylating and crosslinking agents (BCNU, chlorambucil, and thiotepa) and two intercalating agents (adriamycin and antinomycin-D) for polyamine-depleted and control cells. The mechanisms producing chemosensitization will be investigated. First, we will compare the rate of drug entry into control and DFMO-pretreated cells. For alkylating and crosslinking agents, we will measure covalent incorporation of drug into macromolecules of control and polyamine-depleted cells. durg-induced damage to intracellular DNA will be assessed using alkaline elution of DNA from membrane filters to measure strand breaks and crosslinks. The ability of intercalating agents to inhibit DNA and/or RNA synthesis will be compared for polyamine-depleted and control cells. We will also use a series of paired normal and malignant cell lines to compae chemosensitization by DFMO pretreatment in tumor and non-tumor cells. To study the function of polyamines in the mammalian cell cycle, cells will be synchronized after DFMO pretreatment by centrifugal elutriation. Cell cycle distributions will be determined by flow cytometry. Synchronized cells will be returned to culture in the presence and absence of exogenous polyamaines and cell cycle progression monitored by changes in cell cycle distribution. We will use this approach to study both inhibition of the G1 to S transition and increased transit time of other cell cycle phases in polyamine-depleted cells. We will also measure the rate of re-appearance of catalytically active ornithine decarboxylase after replating cells pretreated with DFMO.